In HD mice models, LY2510924 biological activity suggesting the involvement of this miRNA in disease

In HD mice models, LY2510924 biological activity suggesting the involvement of this miRNA in disease progression may be restricted to primates. On the contrary, we found miR-128a downregulated in the HD monkeys and human patients, and there are published reports also showing its corresponding downregulation in at least 2 distinct HD mice models [23]. Additionally, our human data for miR-128a corresponds with other reported datasets in HD patients [24]. The miR-128a results, however, were not previously highlighted in the previous publications. Although it is important to identify both primatespecific and conserved molecular mechanisms in disease, we focused on miR-128a in this study due to the compelling list of genes it is predicted to target in HD, including; SP1, HIP1 as well as HTT itself. Our in vitro luciferase reporter assays revealed the binding sites for miR-128a in HIP1, SP1, and HTT are all regulated by miR-128a compared to stringent site-specific mutant controls. Although we experimentally examined a subset of the predicted miR-128a gene targets, there are likely more genes which miR-128a regulates within the HD pathway. Furthermore, a few of the other 11 miRNAs weKocerha et al. Molecular Brain 2014, 7:46 http://www.molecularbrain.com/content/7/1/Page 5 ofFigure 3 Neuropathology in HD monkey frontal cortex. A-F) An increased number of caspase-3 positive cells in frontal cortex of HD monkeys (D-E) was observed when compared with control monkeys (A-B). C) Quantification of caspase-3 positive cells in HD and control monkey frontal cortex showed significant difference between HD monkeys (HD7 and HD8) and control monkey (C1 or C3). However, no difference was observed between HD4 and the controls. The data are presented PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/25746230 as mean ?SE. *P < 0.05 compared with C1, and #P < 0.05 compared with C3. G-L) An increased number and intensity of GFAP positive cells was observed in HD monkeys (HD7 and HD8) but not in HD4 (J-L). High magnification (inserts) showed reactive astrocytes in HD7 and HD8 with strong GFAP staining. Quantification of GFAP expression in the HD monkey frontal cortex showed significant increase expression of GFAP in HD7 and HD8 but not in HD4. The data were presented as mean ?SE. *P < 0.identified to be associated with HD in this study are also likely to regulate genes within the HD pathway. A future point of interest could examine whether a distinct group of miRNAs work together as a network to govern the HD genes. MiR-940, one of the 11 HD-associated miRNAs in the transgenic monkeys, is also predicted totarget HTT. Correspondingly, although we examined miR-128a regulation of the SP1 transcription factor, other miRNAS identified in this study also bioinformatically bind to SP1 (such as miR-320, miR-133, and miR181). Overall, this report points to a collective miRNA targeting of HTT or genes which regulate HTT (i.e. SP1,Kocerha et al. Molecular Brain 2014, 7:46 http://www.molecularbrain.com/content/7/1/Page 6 ofFigure 4 In-vitro analysis of HD-associated miR-128a gene targets. The regulation of the 3'-UTR activity by miR-128 for (A) HIP-1, (B) HTT, (C) SP-1, and (D) GRM-5 was evaluated in 293-FT cells. 293-FT cells were co-transfected with 10 nM pre-miR-128a or negative control (NC) as well as luciferase reporter constructs containing the WT or mutant (MUT) 3'- predicted miR-128a binding site for each of the genes. Relative luciferase activity data were normalized as a percentage of plasmid-only control levels (100 ) for each 3'-UTR construct. One-way.